Bottom Line:
The present study is aimed at deciphering how the mutant IDH can affect cancer pathogenesis, in particular with respect to its associated oncometabolite D-2HG.However, although mutant IDH reduced cell sensitivity to the apoptotic inducer etoposide, D-2HG exhibited no effect on apoptosis.Instead, we found that the apoptotic effect was mediated through the mitochondrial NADH pool reduction and could be inhibited by oxamate.

ABSTRACTSomatic mutations in isocitrate dehydrogenase (IDH)-1 and -2 have recently been described in glioma. This mutation leads to a neomorphic enzymatic activity as the conversion of isocitrate to alpha ketoglutarate (αKG) is replaced by the conversion of αKG to D-2-hydroxyglutarate (D-2HG) with NADPH oxidation. It has been suggested that this oncometabolite D-2HG via inhibition of αKG-dioxygenases is involved in multiple functions such as epigenetic modifications or hypoxia responses. The present study is aimed at deciphering how the mutant IDH can affect cancer pathogenesis, in particular with respect to its associated oncometabolite D-2HG. We show that the overexpression of mutant IDH in glioma cells or treatment with D-2HG triggered an increase in cell proliferation. However, although mutant IDH reduced cell sensitivity to the apoptotic inducer etoposide, D-2HG exhibited no effect on apoptosis. Instead, we found that the apoptotic effect was mediated through the mitochondrial NADH pool reduction and could be inhibited by oxamate. These data show that besides D-2HG production, mutant IDH affects other crucial metabolite pools. These observations lead to a better understanding of the biology of IDH mutations in gliomas and their response to therapy.

Mentions:
Interestingly, our data show that both mutant IDH1 and mutant IDH2 that, respectively, are expressed in cytosol and mitochondria, lead to decreased mitochondrial respiratory reserve. Numerous studies have shown that IDH mutants use αKG as a substrate to produce D-2HG instead of producing it. This drain of αKG must be balanced to some extent by the cells. There are two ways of converting glutamate (Glu) into αKG, either by deamination through glutamate dehydrogenase (GDH) or by transamination through aspartate aminotransferase (AAT).28 Usually, AAT functions in tandem with the malate dehydrogenase (MDH) in the malate–aspartate shuttle (MAS), which transfers reducing equivalent NADH from the cytosol to mitochondria. Glycolysis, which is highly upregulated in cancer cells, is a key source of the reduced form of cytosolic NADH, mainly at the level of LDH.29 As a result, the activity of MAS is increased to shuttle the glycolytic NADH into mitochondria.30 However, it has also been shown that the presence or the lack of mitochondrial substrates could greatly influence the ability of AAT to effectively compete with GDH for Glu.31 We speculate that to limit cytosolic and mitochondrial αKG depletion induced by the presence of mutant IDH, cells are using AAT, independently of MDH, to produce αKG rather than to shuttle NADH in mitochondria (Figure 7). As a result, the dissociation of AAT activity from MDH results in the accumulation of the reductive power of malate trapped in the cytosol and the reduction of mitochondrial NADH pool built from the MAS activity. Altogether, these effects lead ultimately to a reduced mitochondrial respiratory reserve. Further experiments such as direct inhibition of AAT need to be performed in order to confirm our hypothesis. However, it is in agreement with metabolomic studies showing that mutant IDH is associated with decreased fumarate and malate levels, as well as mitochondrial dysfunction.32 Furthermore, it is reinforced with the recovery of the mitochondrial respiratory reserve with oxamate. Indeed, besides being an inhibitor of LDH, oxamate also inhibits AAT.33, 34 In our study, oxamate did not decrease lactate secretion as predicted by its LDH inhibitory action but instead prevented the mitochondrial respiratory reserve loss only in mutant IDH-overexpressing cells.

Mentions:
Interestingly, our data show that both mutant IDH1 and mutant IDH2 that, respectively, are expressed in cytosol and mitochondria, lead to decreased mitochondrial respiratory reserve. Numerous studies have shown that IDH mutants use αKG as a substrate to produce D-2HG instead of producing it. This drain of αKG must be balanced to some extent by the cells. There are two ways of converting glutamate (Glu) into αKG, either by deamination through glutamate dehydrogenase (GDH) or by transamination through aspartate aminotransferase (AAT).28 Usually, AAT functions in tandem with the malate dehydrogenase (MDH) in the malate–aspartate shuttle (MAS), which transfers reducing equivalent NADH from the cytosol to mitochondria. Glycolysis, which is highly upregulated in cancer cells, is a key source of the reduced form of cytosolic NADH, mainly at the level of LDH.29 As a result, the activity of MAS is increased to shuttle the glycolytic NADH into mitochondria.30 However, it has also been shown that the presence or the lack of mitochondrial substrates could greatly influence the ability of AAT to effectively compete with GDH for Glu.31 We speculate that to limit cytosolic and mitochondrial αKG depletion induced by the presence of mutant IDH, cells are using AAT, independently of MDH, to produce αKG rather than to shuttle NADH in mitochondria (Figure 7). As a result, the dissociation of AAT activity from MDH results in the accumulation of the reductive power of malate trapped in the cytosol and the reduction of mitochondrial NADH pool built from the MAS activity. Altogether, these effects lead ultimately to a reduced mitochondrial respiratory reserve. Further experiments such as direct inhibition of AAT need to be performed in order to confirm our hypothesis. However, it is in agreement with metabolomic studies showing that mutant IDH is associated with decreased fumarate and malate levels, as well as mitochondrial dysfunction.32 Furthermore, it is reinforced with the recovery of the mitochondrial respiratory reserve with oxamate. Indeed, besides being an inhibitor of LDH, oxamate also inhibits AAT.33, 34 In our study, oxamate did not decrease lactate secretion as predicted by its LDH inhibitory action but instead prevented the mitochondrial respiratory reserve loss only in mutant IDH-overexpressing cells.

Bottom Line:
The present study is aimed at deciphering how the mutant IDH can affect cancer pathogenesis, in particular with respect to its associated oncometabolite D-2HG.However, although mutant IDH reduced cell sensitivity to the apoptotic inducer etoposide, D-2HG exhibited no effect on apoptosis.Instead, we found that the apoptotic effect was mediated through the mitochondrial NADH pool reduction and could be inhibited by oxamate.

ABSTRACTSomatic mutations in isocitrate dehydrogenase (IDH)-1 and -2 have recently been described in glioma. This mutation leads to a neomorphic enzymatic activity as the conversion of isocitrate to alpha ketoglutarate (αKG) is replaced by the conversion of αKG to D-2-hydroxyglutarate (D-2HG) with NADPH oxidation. It has been suggested that this oncometabolite D-2HG via inhibition of αKG-dioxygenases is involved in multiple functions such as epigenetic modifications or hypoxia responses. The present study is aimed at deciphering how the mutant IDH can affect cancer pathogenesis, in particular with respect to its associated oncometabolite D-2HG. We show that the overexpression of mutant IDH in glioma cells or treatment with D-2HG triggered an increase in cell proliferation. However, although mutant IDH reduced cell sensitivity to the apoptotic inducer etoposide, D-2HG exhibited no effect on apoptosis. Instead, we found that the apoptotic effect was mediated through the mitochondrial NADH pool reduction and could be inhibited by oxamate. These data show that besides D-2HG production, mutant IDH affects other crucial metabolite pools. These observations lead to a better understanding of the biology of IDH mutations in gliomas and their response to therapy.